Development of a Fluorescent Probe for Measurement of Singlet Oxygen Scavenging Activity of Flavonoids

A turn-on fluorescent probe, HOCD-RB, for monitoring singlet oxygen (¹O₂) was developed by linking rhodamine B as fluorophore with dimethylhomoocoerdianthrone (HOCD) as ¹O₂ reaction site and fluorescence quencher due to the intramolecular energy transfer (ET) between rhodamine B and HOCD moieties. Upon exposure to ¹O₂ it rapidly forms endoperoxide with HOCD and turns on the fluorescence of rhodamine B by 18-fold. Taking advantage of the HOCD-RB probe that shows fast response, high sensitivity, and selectivity for ¹O₂, it is applied for imaging of endogenous ¹O₂ in living cells and the fluorometric assay for evaluating ¹O₂ quenching activity of selected common flavonoids found in our daily diets. The results show that the ¹O₂ scavenging activity of flavonoids depends on not only the structure of individual flavonoid but also the competitive interactions between mixed flavonoids. The best antioxidant capacity for individual and mixed flavonoids is epigallocatechin gallate and the mixture of catechin gallate with kaempferol, respectively. Overall, this work provided a new tool for detection and imaging of singlet oxygen activity in a biological system as well as an efficient fluorometric assay of ¹O₂ scavenging activity.